The skin is the largest organ of the human body. The skin consists of the epidermal and dermal layers. The epidermis is the outer layer, sitting on and nourished by the thicker dermis. These two layers are approximately 1-2 mm (0.04-0.08 in) thick. The epidermis consists of an outer layer of dead cells, which provides a tough, protective coating, and several layers of rapidly dividing cells called keratinocytes. The dermis contains the blood vessels, nerves, sweat glands, hair follicles, and oil glands. The dermis consists mainly of connective tissue, primarily the protein collagen, which gives the skin its flexibility and provides structural support. Fibroblasts, which make collagen, are the main cell type in the dermis.
Skin protects the body from fluid loss, aids in temperature regulation, and helps prevent disease-causing bacteria or viruses from entering the body. Skin that is damaged extensively by burns or non-healing wounds can compromise the health and well-being of the patient. More than 50,000 people are hospitalized for burn treatment each year in the United States, and 5,500 die. Approximately 4 million people suffer from non-healing wounds, including 1.5 million with venous ulcers and 800,000 with diabetic ulcers, which result in 55,000 amputations per year in the United States.
The treatment of severely wounded skin is complicated because the dermis is not capable of self-restoration. One procedure for treating wounded skin is skin grafting. Most commonly, skin grafting is used in the reconstruction of skin after the surgical removal of cutaneous malignancies. However, skin grafts are also used to cover chronic nonhealing cutaneous ulcers, to replace tissue lost in full-thickness burns, or to restore hair to areas of alopecia.
Skin for grafting can be obtained from another area of the patient's body, called an autograft, if there is enough undamaged skin available, and if the patient is healthy enough to undergo the additional surgery required. Alternatively, skin autografts can be made with a person's own keratinocytes through a culturing process. The use of cultured keratinocytes requires an initial small skin biopsy specimen, approximately three weeks to grow the keratinocytes in culture, and may require the use of additional products to stabilize the cultured epidermal layer once surgically applied. Such products may be obtained from another person (donor skin from cadavers is frozen, stored, and available for use), called an allograft, or from an animal (usually a pig), called a xenograft. Allografts and xenografts may also be directly applied should autograft material be unavailable, however these products provide only temporary covering—they are rejected by the patient's immune system within 7-10 days and must be replaced with an autograft.
Treatments of severely wounded skin with cultured autografts have shortcomings. In comparison to natural skin, cultured skin is very fragile because it has no dermal layer, and is more susceptible to infection because it has only a few layers of poorly differentiated cells. The use of cultured skin autografts is costly and requires lengthy culture times. Additionally, residual feeder layer cells (e.g., murine-derived feeder layers) used to support the in vitro growth of keratinocytes may constitute an impurity within the growing cells, cell lines, or cell-derived products. As a result, there is a strong need for improved methods and compositions to detect residual feeder layer cells, especially proliferating feeder layer cells, within cultured keratinocytes and keratinocyte-based products to ensure purity.